Introduction Acute rejection after kidney transplantation, which occurs in 15-20% of recipients, is diagnosed by means of a kidney biopsy which is an invasive procedure. Here we evaluated a novel, minimally invasive approach to diagnose acute rejection by measuring donor-derived cell free DNA (dd-cfDNA) in plasma of kidney transplant recipients.
Methods In total, 18 kidney transplant recipients were studied. Ten recipients experienced a decline in kidney function and underwent a ‘for cause’ biopsy. Of these ten, seven recipients had a biopsy proven acute rejection (BPAR) within the first 4 months after transplantation, while the other three recipients had an urinary tract infection (UTI). Plasma was collected before transplantation, at 3, 6 and 12 months after transplantation and at moment of biopsy. DNA of all 18 donor-recipient pairs was genotyped for 10 highly variable single nucleotide polymorphism (SNP)s. Digital droplet PCR for at least 2 discriminative SNPs between donor and recipient was performed to quantify the level of dd-cfDNA at the different time points.
Results Two recipients with a severe rejection resulting in graft loss, demonstrated a clear dd-cfDNA signal at the moment of rejection with a ratio of dd-cfDNA to total cfDNA of 14.5% and 4%, respectively. The ratio of dd-cfDNA in the other 5 recipients with BPAR was lower with a median of 0.3% (range 0.09 -1.7%), though these percentages corresponded to a significant number of positive dd-cfDNA droplets (median number of droplets: 17; range 9-31). There was a positive correlation between the number of positive dd-cfDNA droplets and serum creatinine concentrations of the recipients with BPAR (r=0.86, p=0.02). The number of positive dd-cfDNA droplets in recipients without BPAR or UTI was low (1; range 0-12) at all time points (n=29 samples). Two recipients with an UTI demonstrated a ratio of 0.4% and 0.04% dd-cfDNA corresponding with 12 and 6 positive dd-cfDNA droplets, while the third patient who experienced a recurring UTI had high levels of dd-cfDNA with a ratio of 11% dd-cfDNA representing 168 dd-cfDNA droplets.
Conclusion Our first data show that dd-cfDNA is a potential minimally invasive marker for acute rejection after kidney transplantation. Nevertheless, the discriminative capacity of dd-cfDNA as marker for rejection from other causes of graft damage or clinical problems as urinary tract infection needs to be elucidated.